Phenotypic characteristics associated with virulence of clinical isolates from the Sporothrix complex

Abstract

The Sporothrix complex members cause sporotrichosis, a subcutaneous mycosis with a wide spectrum of clinical manifestations. Several specific phenotypic characteristics are associated with virulence in many fungi, but studies in this field involving the Sporothrix complex species are scarce. Melanization, thermotolerance, and production of proteases, catalase, and urease were investigated in 61 S. brasiliensis, one S. globosa, and 10 S. schenckii strains. The S. brasiliensis strains showed a higher expression of melanin and urease compared with S. schenckii. These two species, however, presented similar thermotolerances. Our S. globosa strain had low expression of all studied virulence factors. The relationship between these phenotypes and clinical aspects of sporotrichosis was also evaluated. Strains isolated from patients with spontaneous regression of infection were heavily melanized and produced high urease levels. Melanin was also related to dissemination of internal organs and protease production was associated with HIV-coinfection. A murine sporotrichosis model showed that a S. brasiliensis strain with high expression of virulence factors was able to disseminate and yield a high fungal burden in comparison with a control S. schenckii strain. Our results show that virulence-related phenotypes are variably expressed within the Sporothrix complex species and might be involved in clinical aspects of sporotrichosis.

Figure 1

Production of melanin by members of the Sporothrix complex is dependent on the available nutrients in the microenvironment. (a) Degree of melanization and (b) melanization times of 72 Sporothrix strains in Sabouraud Dextrose Agar (SDA) and minimal medium (MM).

Figure 2

Differential production of melanin by S. brasiliensis and S. schenckii sensu stricto. ((a), (b)) Percent melanized strains and ((c), (d)) melanization times of the two species in ((a), (c)) and Sabouraud Dextrose Agar and in ((b), (d)) minimal medium.

Figure 3

Box-plot diagrams showing similar thermotolerances between S. brasiliensis and S. schenckii sensu stricto observed at 21 days of growth in Sabouraud Dextrose Agar.

Figure 4

Correlation between melanization and thermotolerance in the Sporothrix complex. (a) The growth inhibition of the strains is not dependent on their levels of DHN-melanin. (b) Tricyclazole, a DHN-melanin inhibitor, is able to enhance fungal growth inhibition. (c) L-DOPA and (d) L-tyrosine, precursors for eumelanin and pyomelanin, respectively, are able to reduce growth inhibition in some strains.

Figure 5

Urease production, as measured by the optical density (OD) at 559 nm, of the yeast-form of 72 Sporothrix strains at (a) four and (b) seven days of incubation at 37°C in Christensen's urea broth.

Figure 6

Fungal burden after an experimental infection of C57/Bl6 mice by yeast cells of one strain producing several virulence-related phenotypes (IPEC26449) and one strain with low production of these characteristics (ATCC16345). Two different inoculum sizes were used, 5 × 10⁷ (low) and 1 × 10⁸ (high) Sporothrix yeasts. ^* P < 0.05.